Diagnostic Stuart’s Urea Broth Base (Urea Broth Base)
Intended Use:
Recommended for the identification of bacteria on the basis of urea utilization, specially for the differentiation of Proteus,
Salmonella and Shigella species from clinical and non-clinical species.
Composition**
Ingredients Gms / Litre
Potassium dihydrogen phosphate 9.100
Di-potassium hydrogen phosphate 9.500
Yeast extract 0.100
Phenol red 0.010
Final pH ( at 25°C) 6.8±0.2
**Formula adjusted, standardized to suit performance parameters
Directions
Suspend 18.71 grams in 950 ml purified/distilled water. Heat if necessary to dissolve the medium completely. Sterilize by
autoclaving at 15 lbs pressure (121°C) for 15 minutes. Cool to 45-50°C. Aseptically add 50 ml of sterile U40 Supplement
(FD048). Mix well and distribute in 10 ml amounts into sterile tubes. Principle And Interpretation
Rustigian and Stuart developed Urea Broth (1). This medium is especially recommended for the differentiation of
Proteus species from Salmonella and Shigella species in the enteric infection diagnosis (2), based on urea utilization (3,4).
Gram-negative enteric bacilli are unable to utilize urea because of less nutrients and high buffering capacity of the medium.
Urea Broth becomes alkaline as the utilization of urea by the organisms liberates ammonia during the incubation, indicated
by pink red colour. All urea test media rely on the alkalinity formation and so they are not specific for urease testing. The
utilization of proteins may raise the pH to alkalinity due to protein hydrolysis and excess of amino acids results in falsepositive reaction. A medium without urea serves as negative control to rule out false positive results.
Type of specimen
Pure isolate
Specimen Collection and Handling
For food and dairy samples, follow appropriate techniques for sample collection and processing as per guidelines (5,6,7).
For water samples, follow appropriate techniques for sample collection, processing as per guidelines and local standards (8).
For clinical samples follow appropriate techniques for handling specimens as per established guidelines (9,10).
After use, contaminated materials must be sterilized by autoclaving before discarding.
Warning and Precautions
In Vitro diagnostic use. For professional use only. Read the label before opening the container. Wear protective
gloves/protective clothing/eye protection/face protection. Follow good microbiological lab practices while handling
specimens and culture. Standard precautions as per established guidelines should be followed while handling clinical
specimens. Safety guidelines may be referred in individual safety data sheets.
Limitations
1. Prolonged incubation may cause alkaline reaction in the medium.
2. Also, all urea test media rely on the alkalinity formation and so they are not specific for determining the absolute rate of
urease activity (4).
Performance and Evaluation
Performance of the medium is expected when used as per the direction on the label within the expiry period when stored at
recommended temperature. Quality Control
Appearance
Light yellow to light pink homogeneous free flowing powder
Colour and Clarity of prepared medium
Yellowish orange coloured clear solution in tubes.
Reaction
Reaction of basal medium (1.87gm in 95ml distilled water) at 25°C. pH : 6.8±0.2
pH
6.60-7.00
Cultural Response
Cultural characteristics observed on addition of sterile U40 Supplement (FD048) after an incubation at 35-37°C for 18-24
hours.
Organism Inoculum
(CFU)
Urease
# Klebsiella aerogenes
ATCC 13048 (00175*)
50-100 negative reaction,
no change
Escherichia coli
ATCC 8739 (00012*)
50-100
Klebsiella pneumoniae
ATCC 10031
50-100
Escherichia coli
NCTC 9002
50-100
Escherichia coli
ATCC 25922 (00013*)
50-100
ATCC 13883 (00097*)
50-100
50-100
Salmonella Typhimurium
ATCC 14028 (00031*)
Klebsiella pneumoniae
50-100
Proteus vulgaris
ATCC 13315
Proteus mirabilis
ATCC 25933
50-100
negative reaction, no
change
positive reaction,
cerise colour
negative reaction,
no change
negative reaction,
no change
negative reaction, no
change
positive reaction,
cerise colour
positive reaction,
cerise colour
positive reaction,
cerise colour
Key : (*) - Corresponding WDCM numbers.
(#) - Formerly known as Enterobacter aerogenes
Storage and Shelf Life
Store between 10-30°C in a tightly closed container and the prepared medium at 2 - 8°C. Use before expiry date on the label.
On opening, product should be properly stored dry, after tightly capping the bottle in order to prevent lump formation due to
the hygroscopic nature of the product. Improper storage of the product may lead to lump formation. Store in
dry ventilated area protected from extremes of temperature and sources of ignition Seal the container tightly
after use. Product performance is best if used within stated expiry period.
Disposal
User must ensure safe disposal by autoclaving and/or incineration of used or unusable preparations of this
product. Follow established laboratory procedures in disposing of infectious materials and material that comes into
contact with clinical sample must be decontaminated and disposed of in accordance with current laboratory techniques (9,10).
